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BT238

CRL-3414

Product category
Human cells
Organism
Homo sapiens, human
Morphology
irregular aggregates
Tissue
Brain
Disease
Glioblastoma
Applications
3D cell culture
Cancer research
Neuroscience
Product format
Frozen
Storage conditions
Vapor phase of liquid nitrogen
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Price: $664.00 ea
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Documentation

Biosafety Icon BSL 1

ATCC determines the biosafety level of a material based on our risk assessment as guided by the current edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL), U.S. Department of Health and Human Services. It is your responsibility to understand the hazards associated with the material per your organization’s policies and procedures as well as any other applicable regulations as enforced by your local or national agencies.

ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.

Required Products

These products are vital for the proper use of this item and have been confirmed as effective in supporting functionality. If you use alternative products, the quality and effectiveness of the item may be affected.

Detailed product information

Characteristics

Cells per vial
Approximately 2.0 to 3.0 x 106
Volume
1.0 mL
Growth properties
Mixed: suspension and aggregate
Age
61 years
Gender
Male
Comments
MGMT promoter methylation Methylated, EGFR wild-type, PTEN heterozygous mutant, TP53 wild-type, IDH wild-type

Handling information

Unpacking and storage instructions
  1. Check all containers for leakage or breakage.
  2. Remove the frozen cells from the dry ice packaging and immediately place the cells at a temperature below ­-130°C, preferably in liquid nitrogen vapor, until ready for use.
Complete medium
NeuroCult Basal Medium w/ proliferation Supplement + 20 ng/mL EGF + 20 ng/mL β-FGF + 2 μg/mL Heparin sulphate. Protect from light and use complete medium within 10 days.

NeuroCult NS-A Proliferation Kit (Stem Cell Technologies cat# 5751)

  • 450 mL NeuroCult Basal Medium (SCT cat# 5750)
  • 50 mL NeuroCult Supplement (SCT cat# 5753)

20 μg/mL EGF: use 1 μL/1 mL culture medium. To prepare 20 μg/mL EGF stock solution, aseptically combine:

  • 100 μg EGF (PeproTech cat# 100-15)
  • 1 mL PBS (ATCC 30-2200)

Then further dilute by adding 4 mL PBS/0.1% BSA

20 μg/mL β-FGF: use 1 μL/1 mL culture medium. To prepare 20 μg/mL EGF stock solution, aseptically combine:

  • β-FGF (PeproTech cat# 100-18B)
  • 1 mL 5 mM Tris PH 7.6

Then further dilute by adding 4 mL PBS/0.1% BSA

Store in working aliquots at -20°C. EGF and β-FGF is stable for 1 year when prepared and stored as directed. Do not repeat freeze thaw.

2 mg/mL Heparin Solution (Stem Cell Technologies cat# 07980): use 1 μL/1 mL culture medium. Store solution at 4°C.

Temperature
37°C
Atmosphere
95% Air, 5% CO2
Handling procedure
To insure the highest level of viability, thaw the vial and initiate the culture as soon as possible upon receipt. If upon arrival, continued storage of the frozen culture is necessary, it should be stored in liquid nitrogen vapor phase and not at -70°C.  Storage at -70°C will result in loss of viability.

  1. Thaw the vial by gentle agitation in a 37°C water bath. To reduce the possibility of contamination, keep the O-ring and cap out of the water.  Thawing should be rapid (approximately 2 minutes).
  2. Remove the vial from the water bath as soon as the contents are thawed, and decontaminate by dipping in or spraying with 70% ethanol. All of the operations from this point on should be carried out under strict aseptic conditions.
  3. Transfer the vial contents to a centrifuge tube containing 9.0 mL complete culture medium and spin at approximately 125 x g for 5 to 7 minutes.
  4. Resuspend cell pellet with the recommended complete medium (see the specific batch information for the culture recommended dilution ratio) and dispense into a 25 cm2 or a 75 cm2 culture flask.  It is important to avoid excessive alkalinity of the medium during recovery of the cells.  It is suggested that, prior to the addition of the vial contents, the culture vessel containing the complete growth medium be placed into the incubator for at least 15 minutes to allow the medium to reach its normal pH (7.0 to 7.6).
  5. Incubate the culture at 37°C in a suitable incubator.  A 5% CO2 in air atmosphere is recommended if using the medium described on this product sheet.
Handling notes
Neurospheres must be mechanically broken up when they get too large (they form ragged edges and dark areas within the neurospheres). Do not dispense cells into single cell suspension during this process, but just reduce in size of clusters to maintain a healthy state.
Subculturing procedure
Cultures can be maintained by addition or replacement of fresh medium. Start cultures at 2.0 X 105 cells/mL and maintain between 1.0 X 105 and 3.0 X 106 cells/mL.

Medium Renewal: Add fresh medium every 2 to 3 days (depending on cell density).
Reagents for cryopreservation
Complete culture medium (with or without EGF/β-FGF/Heparin) + 10% DMSO (ATCC 4-X). Store in liquid nitrogen vapor phase.

Quality control specifications

Bacterial and fungal testing
Not detected
Mycoplasma contamination
Not detected
Virus testing
Hepatitis B virus (HBV): Not detected
Cytomegalovirus (CMV): Not detected
Human Immunodeficiency virus (HIV): Not detected
Epstein-Barr virus (EBV): Not detected
Human papillomavirus (HPV): Not detected
STR profiling
Amelogenin: X,Y
CSF1PO: 11
D13S317: 8
D16S539: 11,14
D5S818: 12,13
D7S820: 10,12
THO1: 6,9
TPOX: 8
vWA: 16,19

History

Depositors
University of Calgary
Year of origin
2012

Legal disclaimers

Intended use
This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use.
Warranty

The product is provided 'AS IS' and the viability of ATCC® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid.  Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.

Disclaimers

This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use. Any proposed commercial use is prohibited without a license from ATCC.

While ATCC uses reasonable efforts to include accurate and up-to-date information on this product sheet, ATCC makes no warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. ATCC does not warrant that such information has been confirmed to be accurate or complete and the customer bears the sole responsibility of confirming the accuracy and completeness of any such information.

This product is sent on the condition that the customer is responsible for and assumes all risk and responsibility in connection with the receipt, handling, storage, disposal, and use of the ATCC product including without limitation taking all appropriate safety and handling precautions to minimize health or environmental risk. As a condition of receiving the material, the customer agrees that any activity undertaken with the ATCC product and any progeny or modifications will be conducted in compliance with all applicable laws, regulations, and guidelines. This product is provided 'AS IS' with no representations or warranties whatsoever except as expressly set forth herein and in no event shall ATCC, its parents, subsidiaries, directors, officers, agents, employees, assigns, successors, and affiliates be liable for indirect, special, incidental, or consequential damages of any kind in connection with or arising out of the customer's use of the product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of such materials.

Please see the material transfer agreement (MTA) for further details regarding the use of this product. The MTA is available at www.atcc.org.

Permits & Restrictions

Import Permit for the State of Hawaii

If shipping to the U.S. state of Hawaii, you must provide either an import permit or documentation stating that an import permit is not required. We cannot ship this item until we receive this documentation. Contact the Hawaii Department of Agriculture (HDOA), Plant Industry Division, Plant Quarantine Branch to determine if an import permit is required.

MORE INFORMATION ABOUT PERMITS AND RESTRICTIONS

References

Curated Citations

Cusulin C, et al. Precursor States of Brain Tumor Initiating Cell Lines Are Predictive of Survival in Xenografts and Associated with Glioblastoma Subtypes. Stem Cell Reports 5(1): 1-9, 2015. PubMed: 26095605

Kelly JJ, et al. Proliferation of human glioblastoma stem cells occurs independently of exogenous mitogens. Stem Cells 27(8): 1722-33, 2009. PubMed: 19544433

Luchman A, et al. Dual mTORC1/2 Blockade Inhibits Glioblastoma Brain Tumor Intiating Cells In Vitro and In Vivo and Synergizes with Temozolomide to Increase Orthotopic Xenograft Survival. Clin Cancer Res 20(22): 5756-67, 2014. PubMed: 25316808

Stechishin O, et al. On-target JAK2/STAT3 inhibition slows disease progression in orthotopic xenografts of human glioblastoma brain tumor stem cells. Neuro Oncol 15(2): 198-207, 2013. PubMed: 23262510

Frequently Asked Questions

Need assistance with this product? Contact our Technical Support team.

Telephone

Telephone

US and Puerto Rico

800-638-6597

Outside the US

+1-703-365-2700
hours

Hours of Operation

Monday-Friday
9:30am - 5:30pm
US Eastern Time